Dominant Negative Regulation by c-Jun of Transcription of the Uncoupling Protein-1 Gene through a Proximal cAMP-Regulatory Element: A Mechanism for Repressing Basal and Norepinephrine-Induced Expression of the Gene before Brown Adipocyte Differentiation

Abstract

The brown fat uncoupling protein-1 (ucp-1) gene is regulated by the sympathetic nervous system, and its transcription is stimulated by norepinephrine, mainly through cAMP-mediated pathways. Overexpression of the catalytic subunit of protein kinase A stimulated a chloramphenicol acetyltransferase expression vector driven by the 4.5-kb 5'-region of the rat ucp-1 gene. Mutant deletion analysis indicated the presence of the main cAMP-regulatory element (CRE) in the proximal region between -141 and -54. This region contains an element at -139/-122 able to confer enhancer and protein kinase A (PKA)-dependent activity to the basal thymidine kinase promoter. The potency of this element was much higher in differentiated than in nondifferentiated brown adipocytes. Gel shift analyses indicated that a complex array of proteins from brown fat nuclei bind to the -139/-122 element, among which CRE-binding protein (CREB) and Jun proteins were identified. In transfected brown adipocytes, c-Jun was a negative regulator of basal and PKA-induced transcription from the ucp-1 promoter acting through this proximal CRE region. A double-point mutation in the -139/-122 element abolished both PKA- and c-Jun-dependent regulation through this site, and overexpression of CREB blocked c-Jun repression. Thus, an opposite action of these two transcription factors on the -139/-122 CRE is proposed. c-Jun content in brown adipocytes differentiating in culture correlated negatively with both ucp-1 gene expression and the acquisition of the brown adipocyte morphology. These findings indicate that c-Jun provides a molecular mechanism to repress the basal and cAMP-mediated expression of the ucp-1 gene before the differentiation of the brown adipocyte.

p38 Mitogen-Activated Protein Kinase Is the Central Regulator of Cyclic AMP-Dependent Transcription of the Brown Fat Uncoupling Protein 1 Gene

“…Finally, using ChIP studies, we provide evidence that CREB indeed contributes to the cAMP responsiveness of the UCP1 gene by binding the CRE (presumably CRE4) at the proximal region of the promoter. This observation is consistent with a previous report that CREB can interact with the equivalent region of the rat UCP1 promoter (45). Many key features of the brown fat thermogenic response have been shown to be dependent on ␤-adrenergic stimulation of cAMP production, including proliferation (3), UCP1 and type II deiodinase expression, mitochondrial biogenesis, and respiration (27).…”

“…However, there has never been a clearly demonstrated role for CRE-binding protein (CREB) to regulate the UCP1 enhancer, although CREB can bind to the CRE in the proximal region of the rat UCP1 promoter (positions Ϫ139 to Ϫ122). This latter element alone is unable to activate the UCP1 gene but serves to augment the critical role of the UCP1 enhancer (45). Therefore, to date the mechanism(s) whereby cAMP signaling and nuclear factors intersect to coordinate the regulation of UCP1 expression is still unclear.…”

Insulin increases the adrenergic stimulation of 5′ deiodinase activity and mRNA expression in rat brown adipocytes; role of MAPK and PI3K

“…We tried to inhibit proteasome activity with several proteasome inhibitors (such as PSI and MG132). Unfortunately, most inhibitors of proteasome (PSI, MG132 and lactacystin) strongly induce c-Jun kinase (JNK-1) (Meriin et al 1998), a pathway that inhibits adrenergic stimulation by interaction with the cAMP response element (CRE) element (Yubero et al 1998). We did time-course and dose-response experiments with both MG132 and PSI, and the results were similar with both (not shown).…”

Growth factor regulation of uncoupling protein-1 mRNA expression in brown adipocytes

“…Our results confirm EGF as an important growth factor for adipose cells and suggest that EGF, in addition to vasopressin, could play an important role interfering in the process of brown adipocyte differentiation. In this respect, high levels of c-Jun, present in nondifferentiated brown adipocytes as compared with differentiated cells, have been reported to repress the basal and cAMP-mediated expression of UCP-1 gene (36). These authors postulate that c-Jun levels could determine the cellular state, e.g., proliferative or differentiated in brown adipocytes.…”